Spectrophotometer

ABSTRACT

A spectrophotometer including a light source (1) operative to emit a beam of light (15), an optical system for directing the light beam (15) to a sample (8) to be analyzed, and a detector (9) which detects the intensity of the light beam after the beam interacts with the sample (8). The light source (1) is operative to emit bursts of light separated by an interval during which no light is emitted. By way of example, a xenon tube may be used for that purpose. The spectrophotometer measures the intensity of the light beam generated by each burst of light after that beam interacts with the sample. Each such light beam may be divided into first and second parts (5 and 4) prior to interaction with the sample (8), and the optical system is arranged to direct the first part (5) to the sample (8) and to direct the second part (4) to a second detector (7) for conducting a reference measurement. A dark signal measurement may be conducted immediately before or after each burst of light.

This invention relates to spectroscopy and is particularly concernedwith ultraviolet/visible/infrared spectrophotometers.

Conventional ultraviolet/visible/infrared spectrophotometers use one ormore light sources that continuously emit light. For example, a popularcombination is a deuterium arc lamp and a quartz halogen filament lampto cover the ultraviolet and visible/infrared portions of the spectrumrespectively. In order to obtain high quality readings from aninstrument, it is necessary to obtain three separate measurements. Ameasurement of the intensity of the light source without any samplepresent (called the reference measurement), a measurement of theintensity when the sample is present (called the sample measurement) anda measurement of the signal when no light from the source reaches thedetector (called the dark signal). A measurement of the fraction oflight not absorbed by the sample can then be computed as:

(sample-dark)/(reference-dark).

In order to obtain such measurements it is common practice to use amechanical chopping mechanism which, depending on it's position, directsthe beam from the source along either of two paths. One path bypassesthe sample and goes to a detector for reference measurement whereas theother path passes to the sample and then goes to a detector for samplemeasurement. The chopping mechanism also functions to block the lightbeam from the source to the detector for dark measurement.

Such an arrangement has some significant disadvantages. Firstly, sincethe chopper is a mechanical device there is a practical limit at whichthe system can switch from one measurement to the next measurement(typically more than 1 millisecond and frequently about 10milliseconds). Any changes in the system conditions during thisswitching time will not be correctly eliminated. For example, if thelight source fluctuates in output with time, changes in effectiveintensity between sample and reference measurements will not beeliminated. Furthermore, a major component of the dark measurement isroom light entering the optical path between light and detector, and theintensity of room light can fluctuate significantly and rapidly withtime especially if fluorescent lights are used in the room. Changes inthis intensity between dark and sample or dark and referencemeasurements will not be cancelled correctly. The problem isparticularly evident in spectrophotometers in which the spectralresolution subsystem is placed before the sample. In such arrangementsonly a very small fraction of the source light passes through thespectral resolution subsystem and consequently the source lightintensity on the sample is relatively low. Room light ingress istherefore a significant factor and in a conventional instrument it isnecessary to take steps to rigidly exclude all room light.

The aforementioned problem is not so evident in spectrophotometers inwhich the spectral resolution subsystem is placed after the sample andthus illuminate the sample with white light since the total lightintensity from the source is usually much larger than the room lightaccepted into the optical path. On the other hand, such instruments havevery significant limitations when measuring some types of samples. Lightat wavelengths other than the wavelength of interest can excitefluorescence which causes the sample to emit additional light at thewavelength of interest. A second problem is the total amount of lightincident on the sample which can be sufficient to affect the sample andthereby cause errors in the measurement process.

Another disadvantage of the conventional arrangement is that the lightsource usually needs some time to stabilise (for the operatingtemperature to equilibrate) and is therefore left on while theinstrument is switched on, regardless of whether or not measurements arebeing made. Sample measurement occurs over a small portion of the timethe instrument is switched on and as a result continuous energisation ofthe light source wastes power and shortens the useful life of the lightsource. In addition the sample remains illuminated during the time it iswithin the sample compartment and as a consequence the total light loadexperienced by the sample can be very significant in some circumstances.This can be a problem for samples which are photosensitive (quite commonfor kinetics samples).

An object of the present invention is to overcome or at least alleviatethe aforementioned disadvantages. It is a further object of theinvention to provide a spectrophotometer which is relatively inexpensiveand which nevertheless enables relatively accurate sample analysis.

A spectrophotometer according to the invention is characterised in thatthe light source is pulsed so that short intense bursts of light aregenerated and no light is emitted between successive bursts. An examplelight source suitable for that purpose is a Xenon flash tube. It ispreferred that the dark signal measurement occurs at a time close to thetime (e.g., within 500 microseconds) at which the sample and referencemeasurements occur. The dark signal measurement may be effected byexplicitly measuring the signal level directly before a pulse or burstof light is generated, or it may be effected by electronically adjustingthe detector output to zero directly before a pulse or burst of light isgenerated. An instrument according to the invention preferably includestwo detectors and an optical system arranged to permit simultaneousmeasurement of the sample and the reference. At least the samplemeasurement and the reference measurement can be conducted on twooptical signals derived from the same burst of light, and it ispreferred that those signals are simultaneously derived from that burstof light.

Embodiments of the invention are described in detail in the followingpassages of the specification which refer to the accompanying drawings.The drawings, however, are merely illustrative of how the inventionmight be put into effect, so that the specific form and arrangement ofthe various features as shown is not to be understood as limiting on theinvention.

In the drawings:

FIG. 1 is a diagrammatic illustration of a spectrophotometer accordingto one embodiment of the invention.

FIG. 2 is a diagrammatic illustration of one form of controlled circuitfor use with the spectrophotometer of FIG. 1.

FIG. 3 is a diagrammatic illustration of the manner of operation of thecircuit shown by FIG. 2.

FIG. 4 is a diagrammatic illustration of the relationship between themonochromator entrance slit and the arc position of the light sourcewhich exists in prior art arrangements.

FIG. 5 is a diagrammatic illustration similar to FIG. 4, but showing adifferent relationship as adopted in one embodiment of the invention.

FIG. 6 is a diagrammatic illustration similar to FIG. 1 but showinganother embodiment of the invention.

In the arrangement shown by FIG. 1 a pulsed light source 1 such as aXenon flash tube is operable in a known manner to emit very shortintense bursts of light each of which could, for example, have aduration of 2 to 20 microseconds. No light is emitted in the intervalbetween successive bursts. The optical arrangement shown by FIG. 1includes a fixed beam splitter 2 which divides the incident light beam 3received from the lamp 1 into two beams 4 and 5 having a predeterminedintensity ratio. By way of example, the two beams 4 and 5 can be ofsubstantially equal intensity. The beam 4 is directed through anoptional reference cell 6 and from there passes to a reference detector7. The beam 5 is directed to a sample cell 8 and from there passes to asample detector 9. Such an arrangement has the advantage of enablingsimultaneous detection of the optical signals upon which the referenceand sample measurements are based, thereby eliminating problems whichcan arise out of unpredictable variation in the intensity of the twosignals. The final processing of each of the signals which results inthe reference measurement and the sample measurement respectively, mayor may not occur in parallel at the same time.

The arrangement shown by FIG. 1 involves transmission measurement of thesample. That is, the beam 5 passes through a cell 8 containing thesample to be analysed. It is to be understood that the invention is alsoapplicable to spectrophotometers in which the sample to be analysed issubjected to reflectance measurement and consequently do not require thepresence of a cell as such.

Since the lamp 1 does not emit light in the time interval betweensuccessive bursts of light there is no need for mechanical means tointerrupt the light beam in order to determine the dark signalmeasurement. Furthermore, since the duration of each light pulse isshort it is possible to achieve a very short delay between the darksignal measurement and the sample/reference measurement if the darksignal measurement is effected directly or immediately before or aftergeneration of a lamp pulse. By way of example the delay could be in theorder of 20 to 30 microseconds.

The dark signal measurement could be effected by explicitly measuringthe signal level immediately before the lamp 1 is triggered to generatea pulse of light, and subtracting that measurement from other readingsas adopted in conventional systems. Alternatively, the dark signalmeasurement could be effected by electronically adjusting the detectoroutput to zero immediately before the lamp 1 is triggered to generate apulse of light. An example arrangement utilising that alternative isillustrated by FIGS. 2 and 3.

FIG. 2 shows an example circuit layout in which the signal from thesample detector 9 passes through an amplifier 10 and depending upon thecondition of switches as hereinafter discussed travels from theamplifier 10 through a circuit including a buffer 11, integrator 12 anda further buffer 13. A corresponding circuit layout will be provided forthe signal generated by the reference detector 7, but is not shown forconvenience of illustration. The following description of the circuitlayout of FIG. 2 is therefore to be understood as also applying to thecorresponding circuit layout for the reference detector 7.

During the time preceding the light pulse, the switch SW1 is held closedto ensure that the input to the buffer amplifier 11 is held at zero. Anyoutput from the detector amplifier 10 appears as a voltage across thecapacitor C1 where it is automatically subtracted from any subsequentlight reading. At the same time, switch SW2 is opened and switch SW3 isclosed to ensure that the integrator 12 is held reset (at groundpotential). When the light pulse is generated the switches SW1, SW2 andSW3 are opened, closed and opened respectively as shown by FIG. 3, andswitch SW4 is closed a short time (T2) before the light pulse so as toallow the sample/hold amplifier 13 to be reset by the integrator 12. Thecondition of the various switches at that time allows the integrator 12to start integrating the detector signal 9 by way of the amplifier 10and the buffer 11. Only changes in the output of the detector amplifier10 since switch SW1 was opened will appear at the input of the bufferamplifier 11 for the reason described above. After the light pulseceases the switch SW1 is closed and the switch SW2 is opened to preventfurther integration. Switch SW4 remains closed for a short period oftime (T1) to allow the sample/hold buffer 13 to settle after which theswitch SW4 opens and the switch SW3 closes holding the integrated outputfrom the detector 9 on the output of buffer 13.

Circuit layouts other than that described above could be adopted toachieve the same result.

The lamp 1 may have a particular disposition relative to themonochromator 14 (FIG. 1). The arc position of the Xenon flash tubetends to move from flash to flash with the result that the reflectedbeam of light 15 from the lamp 1 will only occasionally be accuratelypositioned over the entrance slit 16 of the monochromator 14. That hasthe effect of causing significant variation in the energy received bythe monochromator from successive flashes of the lamp 1. In aconventional arrangement as shown by FIG. 4, the arc of the lamp 1 isdisposed relative to the monochromator slit 16 such that the arcmovement, which is represented by line 17' compared to line 17, istransverse to the direction of the slit 16. Reference 18 represents theimage of the lamp electrodes. In a possible arrangement according to thepresent invention the lamp arc may be disposed so that arc movement isin the direction of the slit 16 as shown by FIG. 5.

An arrangement as shown by FIG. 5 can reduce the sensitivity to arcmovement, but could reduce the light throughput. If desired, loss oflight might be minimised by using a flash tube having a short arclength. For example, an arc length of 1.5 to 2 millimeters might besuitable.

It will be apparent from the foregoing description that aspectrophotometer incorporating the present invention is relativelyinsensitive to variation in room light because of the brief delaybetween the dark signal measurement and sample/reference measurement. Itis therefore not necessary to place the sample in a light proofcompartment and that has a number of benefits.

A further advantage of a spectrophotometer according to the invention isthat simultaneous detection of sample and reference beams results in asystem largely immune to fluctuations in light source intensity. Use ofa pulsed light source then enables energisation of the light source tobe confined to the time over which a measurement is to be made, therebyreducing power consumption and very significantly extending the life ofthe light source. Furthermore, the rate of pulsing the light source canbe adjusted according to the measurement being made. For example, whencarrying out measurements of sample absorbance versus wavelength thelight source can be pulsed very rapidly (typically about 100 times persecond or even faster if necessary compared to a mechanical choppingrate in conventional instruments of typically 30 times per second). Thisallows many more data points to be collected per second leading tofaster analyses. Alternatively, when conducting lengthy kineticsexperiments the flash rate can be reduced, thus reducing the total lightload on the sample while still collecting enough data to characterisethe variation of absorbance with time. For example, if the duration ofthe kinetics experiment is say 1 hour, a reading every 10 seconds isquite sufficient to determine the absorbance/time relationship. The lampis therefore flashed only once every 10 seconds. Compared to aconventional arrangement, this reduces the total light load on thesample by about a factor of 1000 times (typically the light load imposedby the system on the sample with the lamp flashed 100 times per secondis comparable to conventional instrument). This difference can mean thedifference between an accurate answer and meaningless data.

Another advantage of a spectrophotometer according to the invention isthat the pulsing of the light source (and thus a measurement instant)can be synchronised with an external event or condition. The combinationof a short duration pulse and control of the triggering of a pulse makesit possible to synchronise a measurement instant with an external event(or another event within the measurement system) to within microseconds.For example, as shown diagrammatically by FIG. 6, the invention allowsmeasurements to be taken at particular instants on a continuously movingsystem with considerable accuracy. Such a continuously moving systemcould be a carousel 19 carrying cuvettes 8 each of which contains arespective sample for spectrophotometric analysis. Any suitable controlmeans 20 can be adopted to synchronise positioning of the samplescuvettes 8 with emission of bursts of light by the light source 1. Thusthe invention provides an improved measurement resolution which allowsfor such a carousel 19 to be rotated smoothly and continuously and formeasurements to be taken on each sample as it passes through themeasurement position. In contrast, the measurement resolution in theprior art using a mechanical chopping mechanism requires that such acarousel 19 be rotated in a step-wise manner to bring each sample to themeasurement position and to be held there for sufficient time for themeasurements to be taken. It is believed that the invention can provideup to 3 orders of magnitude improved time resolution for some tasks.

Each of the blocks 21 shown in FIG. 6 represents a circuit of the kinddescribed in connection with FIG. 2.

Still further advantages are realizable from an instrument according tothe invention in that its heat generation is less than that ofconventional prior art instruments Generally, conventional instrumentsuse light sources with a combined heat dissipation close to 100-120watts and a total instrument dissipation typically in excess of 150watts. If the heat generating system of such a prior art instrument ishermetically sealed, complex heat exchange structures must be providedto ensure extraction of this energy, otherwise the instrument will heatup to an unacceptable degree. By contrast, an instrument according tothe invention may have an average lamp dissipation of 1 watt and a totalinstrument dissipation of about 3 watts. It can thus be hermeticallysealed readily without requiring complex cooling systems or riskingexcessive temperature rise. Such hermetic sealing may be necessary foran instrument that is to operate in a corrosive or a dangerousenvironment (such as in the presence of flammable gases). Hermeticsealing may also be necessary to eliminate any possibility of ozoneemission to the atmosphere. Ozone is an inevitable by-product of thegeneration of UV light down to 190 nm. An instrument according to thepresent invention is advantageous in this respect in that its lightsource, by virtue of its intermittent and low power operation, generatesless ozone than conventional sources.

In one embodiment of the invention at least the light source and theoptical system of the spectrophotometer are sealed against substantialingress of corrosive or interfering vapours and gases in the surroundingatmosphere. But it is generally convenient to confine the sealedenvironment to those parts of the optical system other than the part atwhich the sample is located. That may be necessary or appropriate toenable convenient placement of the sample to be analysed.

Various alterations, modifications and/or additions may be introducedinto the constructions and arrangements of parts previously describedwithout departing from the spirit or ambit of the invention as definedby the appended claims.

What is claimed is:
 1. A spectrophotometer including a light sourceoperative to emit bursts of light, each two successive bursts of lightbeing separated by an interval during which no light is emitted by saidlight source, an optical system for directing a beam of each said burstof light to a sample to be analyzed, and a detector which detects theintensity of said light beam after interaction of said beam with saidsample and measures a dark signal in immediate proximity to said burstduring said interval.
 2. A spectrophotometer according to claim 1,wherein said light source is a xenon flash tube.
 3. A spectrophotometeraccording to claim 1 or 2, wherein said detector is operative to conducta dark signal measurement immediately before or after each said burst oflight is emitted.
 4. A spectrophotometer according to claim 3, whereinsaid dark signal measurement is effected by measuring the level of anoutput signal of said detector immediately before or after said burst oflight is emitted.
 5. A spectrophotometer according to claim 3, whereinsaid dark signal measurement is effected by adjusting the output of saiddetector to zero immediately before said burst of light is emitted.
 6. Aspectrophotometer according to claim 5, wherein said detector output isadjusted electronically to zero by means of a circuit including, a firstbuffer for receiving a signal from said detector, a capacitor connectingthe output of said first buffer to a high impedance input of a secondbuffer, and a switch operative to connect said input of said secondbuffer to zero.
 7. A spectrophotometer according to claim 6, whereinsaid detector is a first detector for conducting a sample measurement,and a second detector is provided for conducting a referencemeasurement.
 8. A spectrophotometer according to claim 7, wherein saidoptical system includes a beam splitter which divides said beam intofirst and second beam parts prior to said sample, and said systemdirects said first beam part to said sample and said second beam part tosaid second detector.
 9. A spectrophotometer according to claim 8,wherein a reference cell is located in the path of said second beam partbefore said second detector.
 10. A spectrophotometer according to claim7, wherein said sample measurement and said reference measurement areconducted on two optical signals derived from the same said burst oflight.
 11. A spectrophotometer according to claim 7, wherein a saidcircuit is connected to the output of each said detector.
 12. Aspectrophotometer according to claim 2, wherein said optical systemincludes a monochromator having an entrance slit, and the longitudinalaxis of said slit extends in substantially the same direction as that inwhich the arc position of said xenon flash tube tends to move betweensuccessive said bursts.
 13. A spectrophotometer according to claim 1,including means for carrying at least two said samples and which isoperable to move each said sample in succession into and out of the pathof said light beam, and control means which synchronises operation ofsaid light source with movement of said sample carrying means so thatsaid light beam interacts with a said sample while that sample islocated within said path.
 14. A spectrophotometer according to claim 13,wherein each said sample moves into and out of said light beam pathwithout pause.
 15. A spectrophotometer according to claim 1, wherein atleast said light source and said optical system are sealed againstsubstantial ingress of corrosive or interfering vapours and gases in thesurrounding atmosphere.
 16. A spectrophotomer according to claim 15,wherein said seal does not extend to that part of said optical system atwhich said sample is located.
 17. A method of conducting spectroscopicanalysis of a sample comprising the steps of, generating successivebursts of light by a light source which does not emit light during atime period separating each two successive said bursts of light,directing the light beam generated by each said burst through theentrance slit of a monochromator to the sample to be analyzed andmeasuring a light proportional parameter to obtain a measured spectralvalue, measuring said light proportional parameter during a portion ofsaid time period immediately proximate a corresponding burst to obtain adark correction signal, measuring the intensity of said light beam afterit has interacted with said sample, and reducing said measured value bythe dark correction signal to obtain a corrected light proportionalparameter.
 18. A method according to claim 17, wherein said light beamis divided into first and second parts, directing said first part tosaid sample and from there to a first detector, directing said secondpart to a second detector, and measuring the intensity of both said beamparts for each said burst of light.
 19. A method according to claim 18,wherein said second beam part interacts with a reference cell beforebeing received by said second detector.
 20. A spectrophotometercomprising a light source operative to emit bursts of light, each twosuccessive bursts of light being separated by an interval during whichno light is emitted by said light source, said light source possiblytending to produce said bursts in slight random displacement from alight source axis, an optical system for directing a beam of each saidburst of light to a sample to be analyzed, said optical systemcomprising a monochromator having an entrance slit oriented at asubstantial angle with respect to said light source axis, and a detectorwhich detects the intensity of said light beam after interaction of saidbeam with said sample.